Fibronectins are a group of high molecular weight adhesive glycoproteins present in extracellular matrices and plasma. Fibronectins are produced by many types of cultured cells including fibroblasts, epithelial and endothelial cells, myoblasts, glial cells, and macrophages. Oncogenic transformation of these cells causes a marked decrease in the biosynthesis of fibronectin and its deposition in the extracellular matrix. Such cancer cells exhibit a proclivity toward dislodging from the tissue matrix and migrating in the circulatory system to form distant metastases.
Fibronectins isolated from plasma and from normal cells in culture are very similar in their chemical, biological, and immunological properties, yet there are clear differences between them in subunit size, carbohydrate structure, disulfide-dependent polymerization, solubility, ability to agglutinate sheep erythrocytes, and ability to restore normal fibroblast morphology to transformed cells. Fibronectins produced by transformed cells may also be different from those produced by normal cells. J. Biol. Chem. 256:7671-7677, 1981; J. Biol. Chem. 256:11708-11715, 1981. The basis for the molecular and functional polymorphism of fibronectins is poorly understood.
It is recognized that fibronectins are composed of multiple domains which differ in their biological activities. These domains can be separated by limited proteolysis and subsequent affinity chromatography using different ligands. For example, human plasma fibronectin can be cleaved by mild thermolysin digestion into at least four functionally distinct fragments with M.sub.r =155,000-145,000 (Cell/Hep-2 domain), 44,000 (Gel domain), 24,000 (Hep-1/Fib-1 domain), and 22,000 (Fib-2 domain). J. Biol. Chem. 258:3967-3973, 1983. The domain structure of fibronectins produced by cells in culture is, however, less well-characterized than that of plasma fibronectin.
Since fibronectin is the major component of the pericellular matrix, is greatly reduced upon oncogenic transformation, and displays multifunctional properties, including opsonic activity in plasma, extensive and diversified studies have been developed toward understanding the role of this macromolecule in the structure and function of the pericellular matrix as well as in homeostatis of "milieu interieur." For example, in an extensive comparative study of the domain structure of chicken fibronectins, thermolysin fragments of plasma fibronectin and cellular fibronectin separated by various affinity columns were compared. J. Biol. Chem. 256:11292-11300, 1981. In another study, differences between plasma fibronectin and cellular fibronectin in the COOH-terminal domain were suggested by a monoclonal antibody which reacts preferentially with cellular fibronectin. Cell 25:133-141, 1981.
Also previously, Ruoslahti et al. observed a similarity in molecular weight and isoelectric point between fibronectins from human germ cell tumors and amniotic fluid and a distinctive difference from fibronectin of plasma, and suggested that such fibronectins may provide oncodevelopmental markers. Int. J. Cancer 27:763-767, 1981. A great deal of glycosylation diversity in fibronectin has been found to be dependent on fetal or transformed cell origin. J. Bio. Chem. 259:3962-3970, 1984; J. Biol. Chem. 258:5603-5607, 1983; Glycoconjugate 1:155-169, 1984. Heterogeneity of fibronectin caused by phosphorylation or sulfation may also vary with oncogenic transformation. J. Cell. Biol. 80:784-791, 1979; Proc. Natl. Acad. Sci. USA 82:34-37, 1985.
In another previous study, a highly sensitive and specific procedure for the analysis of the domain structure of fibronectins using a panel of antibodies specific for each functional domain was developed. Biochem. Biophys. Res. Comm. 116(2):534-540, 1983.
That method was very recently applied to three human fibronectins isolated from plasma and from the culture supernatants of normal and transformed fibroblasts, which were cleaved by limited proteolysis with thermolysin, trypsin, and cathepsin D as well as by S-cyanylation, followed by identification of each domain with specific antibodies. The results indicate that the COOH-terminal heparin-binding domain, Hep-2, of cellular fibronectins produced by normal and transformed fibroblasts is significantly larger than the same domain of plasma fibronectin. A significant difference in the fragments derived from the Hep-2 domain and from the gelatin-binding domain between the cellular fibronectins produced by normal and transformed fibroblasts was also demonstrated. J. Biol. Chem. 260(8):5105-5114, Issue of Apr. 25, 1985.